Transformation of a normal cell to malignant derivatives is a multistep process that reflects genetic alterations and defects in normal cell proliferation and homeostasis. A fundamental aspect of cancer is unregulated cell cycle control. Unlike normal cells, which only proliferate in response to developmental or other mitogenic signals which is required for tissue growth, the proliferation of cancer cells proceeds without any regulation. Malignant cells also undergo the same cell cycle stages but the checkpoints of cell cycle remain functionless. Cancer cells proliferate because they are insensitive to growth inhibitory signals arising from the stroma or from gene expression pattern changes consequent to ‘terminal’ differentiation, nor do they necessarily require extrinsic growth factors to maintain their proliferative state. Cancer is a manifestation of alterations in cell physiology that dictate uncontrolled cell proliferation. Characteristics of cancer cells include unresponsiveness to programmed cell death, insensitivity to antigrowth signals, independent of growth stimulatory signals, uncontrolled replicative potential and persistent angiogenesis.
Cancer cells have defects in regulatory mechanism that govern normal cell proliferation and homeostasis. Mutation in p53 tumor suppressor gene is a common incident in many human cancers. In normal cells, low levels of p53 is maintained by Mdm2. Mdm2 directly suppress p53 by unmasking its nuclear export signal and its subsequent degradation in cytosol. Elevation of p53 level occurs in response to cellular stress such as DNA damage and that leads to cell cycle arrest and apoptosis. Upon sensing DNA damage, p53 phosphorylates and stabilizes, where it acts as a transcription factor for target genes like p21 a cyclin-dependent kinase inhibitor, Bax a proapoptotic member of the Bcl2 family of proteins, DNA repair proteins and also its own regulator Mdm2. p53 also triggers apoptosis by activating Bax and Bak proapoptotic proteins [Yee, 2005].
Normal cells require mitogenic growth signals to enter the proliferative stage whereas cancer cells show uncontrolled proliferation. When quiescent cells enter cell cycle, cyclin D1 is induced in response to mitogenic signals and cyclinD1 assemble with their catalytic partners CDK4 and CDK6 as cell cycle progress through G0 to G1 phase. Constitutive activation of cyclinD1 contributes to the oncogenic transformation of cancer cells. p21 blocks cell cycle progression by inhibiting cyclin-CDK complex and mediates the p53-dependent cell cycle G1 phase arrest [Sherr, 1999].
Since major objective of an anti-cancer drug is to induce apoptosis in cancer cells by triggering caspase activity where a cascade of events ultimately leads to the death of cells, discussed herein below are the prior art on these aspects.
During apoptosis, cells undergo morphological changes: the cell shrinks, shows deformities and looses contact with its neighbouring cells. Chromatin condensation takes place near the nuclear membrane, externalization of phosphatidyl serine occurs in the plasma membrane and finally the cell is fragmented into compact membrane-enclosed structures, called apoptotic bodies. The most important mechanism that occurs during apoptosis is the activation of proteolytic enzymes which eventually leads to DNA fragmentation. Multitude of specific protein substrates responsible for the maintenance of integrity and shape of the cytoplasm or other organelles undergo cleavage [Saraste, 2000].
Caspases play a pivotal role during apoptosis by degenerating the cell structure, eg. by the destruction of nuclear lamina. During apoptosis, nuclear laminae are cleaved at a single site by caspases leading to nuclear membrane breakdown and contributing to chromatin condensation. Caspases also play a significant role in cellular reorganization indirectly by cleaving several proteins involved in cytoskeleton regulation. Caspases ultimately induces cell death and helps in balancing cellular homeostasis.
The present study was undertaken to assess the anti-proliferative and apoptotic potentiality of different herbal compounds in lung cancer cells and to determine the underlying molecular mechanism behind apoptotic cell death. Several anti-cancer compounds have the potential to suppress cell proliferation but in the present global scenario where cancer is emerging as the greatest threat to human being, specific active compounds are needed which have precise targets.
The present invention is based on herbal source and is important in the sense that the compounds isolated are volatile and therefore can be inhaled. Further, it is desirable that the ideal molecules would select only malignant cells and would not have any impact on normal cells. It may be noted that there is no report on the synergistic effect of compounds present in the Litsea cubeba seed oils and particularly the three compounds used in the present invention in relation to anti-cancer activities. Also, this is for the first time that the vapor of these three compounds isolated from the seed oil of Litsea cubeba has been shown to possess strong anti cancer activities against four cancer cell lines.
Thus, keeping in view the hitherto reported prior art, it may be summarized that the most important requirement as of date is to provide herbal pharmaceutical compositions having anticancer activity which do not have detrimental effects on other body cells. Further, till date there is no report on synergism in relation to the said oils towards anticancer activity.